Circadian Regulation of Neuropeptides from the Hypothalamus.

下丘脑神经肽的昼夜节律调节。

• 批准号: RGPIN-2018-06144
• 负责人: Belsham, Denise
• 金额: $ 3.06万
• 依托单位: University of Toronto
• 依托单位国家: 加拿大
• 项目类别: Discovery Grants Program - Individual
• 财政年份: 2020
• 资助国家: 加拿大
• 起止时间: 2020-01-01 至 2021-12-31
• 项目状态: 已结题
• 来源: https://www.nserc-crsng.gc.ca/ase-oro/Details-Detailles_eng.asp?id=714083
• 关键词: Circadian; Regulation; Neuropeptides; Hypothalamus.

The central or master biological clock is found in the SCN, located in the hypothalamus. This clock operates at the molecular level through a combination of feedback loops, involving clock genes. At a basic level, hypothalamic nuclei outside the SCN express neuropeptides that are directly responsible for the control of energy balance, including neuropeptide Y (NPY) and proopiomelanocortin (POMC). While the SCN does indeed drive the coordination of overall circadian rhythms within the hypothalamus, we hypothesize that an endogenous cellular circadian system in these efferent neurons is required for the rhythmic expression of key neuropeptides and is essential for the control of energy balance. We also predict that the normal rhythmicity can be altered by nutrient excess and endocrine disrupting chemicals (EDC), classified as “obesogens”. We have used immortalized cell lines that express NPY/AgRP or POMC to demonstrate the circadian expression of these neuropeptides. We have also published that the saturated fatty acid palmitate can disrupt these rhythms, with omega-3 DHA rescuing the altered rhythms. Recently, my laboratory has found that the EDC bisphenol A (BPA) alters the expression of NPY and POMC. We will link changes to the intrinsic neuronal circadian machinery (including the rhythmic expression of clock genes Bmal1 and Per2) to hypothalamic neuropeptide gene expression using newly created hypothalamic cell lines from male and female Bmal1 knockout mice. We will also study how neuroinflammation, induced by palmitate and BPA, affects the normal circadian control of clock genes and neuropeptide expression and delineate the molecular mechanisms involved in this process. Aim 1: Understanding the molecular events involved in the palmitate-mediated disruption of rhythmic or circadian control of NPY/AgRP and POMC. We will define the role of the clock genes in the transcriptional regulation of these key genes using novle Bmal1 knockout cell lines and sophisticated molecular techniques, including siRNA knockdown and detailed promoter analysis. Aim 2: Characterizing the role of cellular neuroinflammation, caused by the saturated fatty acid palmitate or BPA, in the cyclical regulation of clock genes and hypothalamic neuropeptides. Changes in rhythmicity will be analyzed for mechanistic alterations at the levels of signal transduction pathways, epigenetic control, and the gene promoter. Although it is generally accepted that the body operates quite routinely on a classic day/night schedule, the molecular events involved in the circadian regulation of basic physiological processes are not yet understood. Since the intact circadian clock is critical for maintaining metabolic processes, including energy homeostasis in vivo, we now intend to define the hypothalamic neurons that play a role in this process, and ultimately to identify the underlying molecular mechanisms.

中央生物钟或主生物钟位于下丘脑的SCN中。这种时钟通过涉及时钟基因的反馈环的组合在分子水平上运作。在基础水平上，下丘脑SCN外的核团表达直接负责控制能量平衡的神经肽，包括神经肽Y(NPY)和前阿片黑素皮质素(POMC)。虽然SCN确实驱动了下丘脑整体昼夜节律的协调，但我们假设这些传出神经元中的内源性细胞昼夜节律系统是关键神经肽有节奏表达所必需的，并且对于能量平衡的控制是必不可少的。我们还预测，正常的节律性可以被营养过剩和内分泌干扰化学物质(EDC)改变，这些物质被归类为“肥胖原”。 我们已经使用了表达NPY/AgRP或POMC的永生化细胞系来证明这些神经肽的昼夜表达。我们还发表了饱和脂肪酸棕榈酸酯可以扰乱这些节律，omega-3 DHA可以挽救改变的节律。最近，我的实验室发现EDC双酚A(BPA)改变了NPY和POMC的表达。我们将使用新建立的来自雄性和雌性BMal1基因敲除小鼠的下丘脑细胞系，将内在神经元昼夜节律机制的变化(包括时钟基因BMal1和PER2的节律性表达)与下丘脑神经肽基因表达联系起来。我们还将研究棕榈酸酯和双酚A诱导的神经炎症如何影响时钟基因和神经肽表达的正常昼夜控制，并描述参与这一过程的分子机制。 目的1：了解棕榈酸干扰NPY/AgRP和POMC节律或昼夜节律控制的分子事件。我们将使用新的BMal1基因敲除细胞系和复杂的分子技术，包括siRNA敲除和详细的启动子分析，来确定时钟基因在这些关键基因的转录调控中的作用。 目的2：研究饱和脂肪酸棕榈酸酯或BPA引起的细胞神经炎症在时钟基因和下丘脑神经肽循环调节中的作用。节律性的变化将被分析为信号转导途径、表观遗传控制和基因启动子水平上的机械性变化。 虽然人们普遍认为人体按典型的昼夜节律运转，但基本生理过程的昼夜节律所涉及的分子事件尚不清楚。由于完整的生物钟对于维持代谢过程至关重要，包括体内的能量动态平衡，我们现在打算定义在这一过程中发挥作用的下丘脑神经元，并最终确定潜在的分子机制。